The studies proposed in this application have the long-term goal to design a novel bone anabolic treatment. Anti-depressants, depending on their selectivity, target the serotonin transporter (SERT) or the norepinephrine transporter (NET) in neurons. The use of anti-depressants of the serotonin transporter family has been associated with bone loss and increased fracture risk. In contrast, anti-depressants of the NET family, in spite of having weak serotonin transporter selectivity, have not shown such a negative effect on bone and even appear to reduce fracture risk in one clinical study. Our preliminary data demonstrates that genetic blockade of NET in mice provokes a dramatic increase in bone volume, caused by a reduction in bone resorption and an increase in bone formation. Net is expressed in nerve cells where its function is to clear norepinephrine (NE) from the synapse, but surprisingly also in osteoblasts, where its function has been unexplored. We have demonstrated in our previous studies that NE released from sympathetic nerves, via the beta 2 adrenergic receptor (2AR), induces bone loss. Based on these studies, one would expect that mice lacking NET would be characterized by NE accumulation at the synapse and thus bone loss. The opposite phenotype we have found in Net-deficient mice led us to propose three possible mechanisms by which Net deficiency could cause a high bone mass: a) chronic 2AR stimulation in osteoblasts that leads to receptor desensitization and blockade of the anti-osteogenic signal from sympathetic nerves, b) low sympathetic tone caused by stimulation of inhibitory a2AR in the brain, and/or c) a cell-autonomous inhibitory function of NET on osteoblast differentiation or function. Regardless of the mechanism of action, our preliminary results identify NET as a potential molecular target for drug discovery of a novel class of bone anabolic agents and have led us to hypothesize that clinically available selective NET blockers could promote bone gain. Our experimental strategies aim at demonstrating this potential anabolic effect NET blockade and at characterizing the molecular mechanism(s) of action underlying this effect. In this application, we propose: 1) To characterize the cellular and structural bone abnormalities induced by Net-deficiency during development and aging, in order to determine whether the phenotype of Net-deficient mice is caused by the acquisition of a high peak bone mass or resistance to age-associated bone loss; 2) To determine the relative contribution of Net deficiency in osteoblasts and neurons to the high bone mass of Net-deficient mice, using both in vivo and in vitro assays to selectively target each cell lineage; 3) To characterize the effect of selective NET inhibitors on bone remodeling, architecture and mechanical properties in mice, as a proof of concept of the notion that selective NET blockers can promote bone gain.